Purified Water

(Ph. Eur. monograph 0008)

H
2
O 18.02 7732-18-5

Ph Eur


DEFINITION

Water for the preparation of medicines other than those that are required to be both sterile and
apyrogenic, unless otherwise justified and authorised.

PURIFIED WATER IN BULK

PRODUCTION

Purified water in bulk is prepared by distillation, by ion exchange, by reverse osmosis or by any
other suitable method from water that complies with the regulations on water intended for
human consumption laid down by the competent authority.

Purified water in bulk is stored and distributed in conditions designed to prevent growth of
micro-organisms and to avoid any other contamination.

Microbiological monitoring

During production and subsequent storage, appropriate measures are taken to ensure that the
microbial count is adequately controlled and monitored. Appropriate alert and action levels are
set so as to detect adverse trends. Under normal conditions, an appropriate action level is a
microbial count of 100 CFU/mL, determined by filtration through a membrane with a nominal
pore size not greater than 0.45 m, using R2A agar and incubating at 30-35 C for not less
than 5 days. The size of the sample is to be chosen in relation to the expected result.


R2A agar





Yeast extract

0.5 g



Proteose peptone

0.5 g



Casein hydrolysate

0.5 g



Glucose

0.5 g



Starch

0.5 g



Dipotassium hydrogen phosphate

0.3 g



Magnesium sulfate, anhydrous

0.024 g



Sodium pyruvate

0.3 g



Agar

15.0 g



Purified water

to 1000 mL




Adjust the pH so that after sterilisation it is 7.2 0.2. Sterilise by heating in an autoclave at 121
C for 15 min.

Growth promotion of R2A agar:
Preparation of test strains. Use standardised stable suspensions of test strains or prepare
them as stated in Table 0008.-1. Seed lot culture maintenance techniques (seed-lot systems)
are used so that the viable micro-organisms used for inoculation are not more than 5 passages
removed from the original master seed-lot. Grow each of the bacterial strains separately as
described in Table 0008.-1. Use buffered sodium chloride-peptone solution pH 7.0 or
phosphate buffer solution pH 7.2 to make test suspensions. Use the suspensions within 2 h, or
within 24 h if stored at 2-8 C. As an alternative to preparing and then diluting a fresh
suspension of vegetative cells of Bacillus subtilis, a stable spore suspension is prepared and
then an appropriate volume of the spore suspension is used for test inoculation. The stable
spore suspension may be maintained at 2-8 C for a validated period of time.

Growth promotion. Test each batch of ready-prepared medium and each batch of
medium, prepared either from dehydrated medium or from the ingredients described. Inoculate
plates of R2A agar separately with a small number (not more than 100 CFU) of the micro-
organisms indicated in Table 0008.-1. Incubate under the conditions described in the table.
Growth obtained must not differ by a factor greater than 2 from the calculated value for a
standardised inoculum. For a freshly prepared inoculum, growth of the micro-organisms must
be comparable to that obtained with a previously tested and approved batch of medium.





















Total organic carbon or oxidisable substances

Carry out the test for total organic carbon (2.2.44) with a limit of 0.5 mg/L or alternatively the
following test for oxidisable substances: to 100 mL add 10 mL of dilute sulfuric acid R and 0.1
mL of 0.02 M potassium permanganate and boil for 5 min; the solution remains faintly pink.

Conductivity

Determine the conductivity off-line or in-line under the following conditions.


EQUIPMENT

Conductivity cell:
electrodes of a suitable material such as stainless steel;

cell constant: the cell constant is generally certified by the supplier and is subsequently
verified at suitable intervals using a certified reference solution with a conductivity less than
1500 Scm
-1
or by comparison with a cell having a certified cell constant; the cell constant is
confirmed if the value found is within 2 per cent of the certified value, otherwise re-calibration
must be performed.

Conductometer Accuracy of 0.1 Scm
-1
or better at the lowest range.

System calibration (conductivity cell and conductometer):
against one or more suitable certified reference solutions;

accuracy: within 3 per cent of the measured conductivity plus 0.1 Scm
-1
.

Conductometer calibration Calibration is carried out for each range of measurement to be
used, after disconnection of the conductivity cell, using certified precision resistors or equivalent
devices with an uncertainty not greater than 0.1 per cent of the certified value.

If in-line conductivity cells cannot be dismantled, system calibration may be performed against
a calibrated conductivity-measuring instrument with a conductivity cell placed close to the cell
to be calibrated in the water flow.

Temperature measurement Tolerance 2 C.


PROCEDURE

Measure the conductivity without temperature compensation, recording simultaneously the
temperature. Temperature-compensated measurement may be performed after suitable
validation.

The water to be examined meets the requirements if the measured conductivity at the recorded
temperature is not greater than the value in Table 0008.-2.



























For temperatures not listed in Table 0008.-2, calculate the maximal permitted conductivity by
interpolation between the next lower and next higher data points in the table.

Heavy metals

If purified water in bulk complies with the requirement for conductivity prescribed for Water for
injections (0169) in bulk, it is not necessary to carry out the test for heavy metals prescribed
below.

CHARACTERS

Appearance

Clear and colourless liquid.

TESTS

Nitrates

Maximum 0.2 ppm.

Place 5 mL in a test-tube immersed in iced water, add 0.4 mL of a 100 g/L solution of
potassium chloride R, 0.1 mL of diphenylamine solution R and, dropwise with shaking, 5 mL of
nitrogen-free sulfuric acid R. Transfer the tube to a water-bath at 50 C. After 15 min, any blue
colour in the solution is not more intense than that in a reference solution prepared at the same
time in the same manner using a mixture of 4.5 mL of nitrate-free water R and 0.5 mL of nitrate
standard solution (2 ppm NO
3
) R.

Aluminium (2.4.17)

Maximum 10 ppb, if intended for use in the manufacture of dialysis solutions.

Prescribed solution To 400 mL of the water to be examined add 10 mL of acetate buffer
solution pH 6.0 R and 100 mL of distilled water R.

Reference solution Mix 2 mL of aluminium standard solution (2 ppm Al) R, 10 mL of acetate
buffer solution pH 6.0 R and 98 mL of distilled water R.

Blank solution Mix 10 mL of acetate buffer solution pH 6.0 R and 100 mL of distilled water R.

Heavy metals (2.4.8)

Maximum 0.1 ppm.

To 200 mL add 0.15 mL of 0.1 M nitric acid and heat in a glass evaporating dish on a water-
bath until the volume is reduced to 20 mL. 12 mL of the concentrated solution complies with
test A. Prepare the reference solution using 10 mL of lead standard solution (1 ppm Pb) R and
adding 0.075 mL of 0.1 M nitric acid . Prepare the blank solution adding 0.075 mL of 0.1 M
nitric acid .

Bacterial endotoxins (2.6.14)

Less than 0.25 IU/mL, if intended for use in the manufacture of dialysis solutions without a
further appropriate procedure for removal of bacterial endotoxins.

LABELLING

The label states, where applicable, that the substance is suitable for use in the manufacture of
dialysis solutions.

PURIFIED WATER IN CONTAINERS

DEFINITION

Purified water in bulk that has been filled and stored in conditions designed to assure the
required microbiological quality. It is free from any added substances.

CHARACTERS

Appearance

Clear and colourless liquid.

TESTS

It complies with the tests prescribed in the section on Purified water in bulk and with the
following additional tests.

Acidity or alkalinity

To 10 mL, freshly boiled and cooled in a borosilicate glass flask, add 0.05 mL of methyl red
solution R. The solution is not coloured red.

To 10 mL add 0.1 mL of bromothymol blue solution R1. The solution is not coloured blue.

Oxidisable substances

To 100 mL add 10 mL of dilute sulfuric acid R and 0.1 mL of 0.02 M potassium permanganate
and boil for 5 min. The solution remains faintly pink.

Chlorides

To 10 mL add 1 mL of dilute nitric acid R and 0.2 mL of silver nitrate solution R2. The solution
shows no change in appearance for at least 15 min.

Sulfates

To 10 mL add 0.1 mL of dilute hydrochloric acid R and 0.1 mL of barium chloride solution R1.
The solution shows no change in appearance for at least 1 h.

Ammonium

Maximum 0.2 ppm.

To 20 mL add 1 mL of alkaline potassium tetraiodomercurate solution R. After 5 min, examine
the solution down the vertical axis of the tube. The solution is not more intensely coloured than
a standard prepared at the same time by adding 1 mL of alkaline potassium tetraiodomercurate
solution R to a mixture of 4 mL of ammonium standard solution (1 ppm NH
4
) R and 16 mL of
ammonium-free water R.

Calcium and magnesium

To 100 mL add 2 mL of ammonium chloride buffer solution pH 10.0 R, 50 mg of mordant black
11 triturate R and 0.5 mL of 0.01 M sodium edetate. A pure blue colour is produced.

Residue on evaporation

Maximum 0.001 per cent.

Evaporate 100 mL to dryness on a water-bath and dry in an oven at 100-105 C. The residue
weighs a maximum of 1 mg.

Microbial contamination

TAMC: acceptance criterion 10
2
CFU/mL (2.6.12). Use casein soya bean digest agar.

LABELLING

The label states, where applicable, that the substance is suitable for use in the manufacture of
dialysis solutions.


Ph Eur







Highly Purified Water
(Ph. Eur. monograph 1927)

H
2
O 18.02 7732-18-5

Ph Eur


DEFINITION

Water intended for use in the preparation of medicinal products where water of high biological
quality is needed, except where Water for injections (0169) is required.

PRODUCTION

Highly purified water is obtained from water that complies with the regulations on water
intended for human consumption laid down by the competent authority.

Current production methods include, for example, double-pass reverse osmosis coupled with
other suitable techniques such as ultrafiltration and deionisation. Correct operation and
maintenance of the system is essential.

In order to ensure the appropriate quality of the water, validated procedures and in- process
monitoring of the electrical conductivity and regular microbial monitoring are applied.

Highly purified water is stored in bulk and distributed in conditions designed to prevent growth
of micro-organisms and to avoid any other contamination.

Microbiological monitoring

During production and subsequent storage, appropriate measures are taken to ensure that the
microbial count is adequately controlled and monitored. Appropriate alert and action levels are
set so as to detect adverse trends. Under normal conditions, an appropriate action level is a
microbial count of 10 CFU per 100 mL when determined by filtration through a membrane with
a nominal pore size not greater than 0.45 m, using R2A agar, at least 200 mL of highly
purified water and incubating at 30-35 C for not less than 5 days.


R2A agar





Yeast extract

0.5 g



Proteose peptone

0.5 g



Casein hydrolysate

0.5 g



Glucose

0.5 g



Starch

0.5 g



Dipotassium hydrogen phosphate

0.3 g



Magnesium sulfate, anhydrous

0.024 g



Sodium pyruvate

0.3 g



Agar

15.0 g



Purified water

to 1000 mL




Adjust the pH so that after sterilisation it is 7.2 0.2. Sterilise by heating in an autoclave at 121
C for 15 min.

Growth promotion of R2A agar
Preparation of test strains. Use standardised stable suspensions of test strains or prepare
them as stated in Table 1927.-1. Seed lot culture maintenance techniques (seed-lot systems)
are used so that the viable micro-organisms used for inoculation are not more than 5 passages
removed from the original master seed-lot. Grow each of the bacterial strains separately as
described in Table 1927.-1. Use buffered sodium chloride-peptone solution pH 7.0 or
phosphate buffer solution pH 7.2 to make test suspensions. Use the suspensions within 2 h, or
within 24 h if stored at 2-8 C. As an alternative to preparing and then diluting a fresh
suspension of vegetative cells of Bacillus subtilis, a stable spore suspension is prepared and
then an appropriate volume of the spore suspension is used for test inoculation. The stable
spore suspension may be maintained at 2-8 C for a validated period of time.






















Growth promotion. Test each batch of ready-prepared medium and each batch of
medium, prepared either from dehydrated medium or from the ingredients described. Inoculate
plates of R2A agar separately with a small number (not more than 100 CFU) of the micro-
organisms indicated in Table 1927.-1. Incubate under the conditions described in the table.
Growth obtained must not differ by a factor greater than 2 from the calculated value for a
standardised inoculum. For a freshly prepared inoculum, growth of the micro-organisms must
be comparable to that obtained with a previously tested and approved batch of medium.

Total organic carbon (2.2.44)

Maximum 0.5 mg/L.

Conductivity

Determine the conductivity off-line or in-line under the following conditions.


EQUIPMENT

Conductivity cell:
electrodes of a suitable material such as stainless steel;

cell constant: the cell constant is generally certified by the supplier and is subsequently
verified at suitable intervals using a certified reference solution with a conductivity less than
1500 Scm-
1
or by comparison with a cell having a certified cell constant; the cell constant is
confirmed if the value found is within 2 per cent of the certified value, otherwise re-calibration
must be performed.

Conductometer Accuracy of 0.1 Scm
-1
or better at the lowest range.

System calibration (conductivity cell and conductometer):
against one or more suitable certified reference solutions;

accuracy: within 3 per cent of the measured conductivity plus 0.1 Scm
-1
.

Conductometer calibration Calibration is carried out for each range of measurement to be
used, after disconnection of the conductivity cell, using certified precision resistors or equivalent
devices with an uncertainty not greater than 0.1 per cent of the certified value.

If in-line conductivity cells cannot be dismantled, system calibration may be performed against
a calibrated conductivity-measuring instrument with a conductivity cell placed close to the cell
to be calibrated in the water flow.

Temperature measurement Tolerance 2 C.


PROCEDURE

Stage 1
1. Measure the conductivity without temperature compensation, recording simultaneously the
temperature. Temperature-compensated measurement may be performed after suitable
validation.

2. Using Table 1927.-2, find the closest temperature value that is not greater than the
measured temperature. The corresponding conductivity value is the limit at that temperature.

3. If the measured conductivity is not greater than the value in Table 1927.-2, the water to be
examined meets the requirements of the test for conductivity. If the conductivity is higher than
the value in Table 1927.-2, proceed with stage 2.

Stage 2
4. Transfer a sufficient amount of the water to be examined (100 mL or more) to a suitable
container, and stir the test sample. Adjust the temperature, if necessary, and while maintaining
it at 25 1 C, begin vigorously agitating the test sample while periodically observing the
conductivity. When the change in conductivity (due to uptake of atmospheric carbon dioxide) is
less than 0.1 Scm
-1
per 5 min, note the conductivity.

5. If the conductivity is not greater than 2.1 Scm
-1
, the water to be examined meets the
requirements of the test for conductivity. If the conductivity is greater than 2.1 Scm
-1
, proceed
with stage 3.

Stage 3
6. Perform this test within approximately 5 min of the conductivity determination in step 5
under stage 2, while maintaining the sample temperature at 25 1 C. Add a recently prepared
saturated solution of potassium chloride R to the test sample (0.3 mL per 100 mL of the test
sample), and determine the pH (2.2.3) to the nearest 0.1 pH unit.

7. Using Table 1927.-3, determine the conductivity limit at the measured pH value in step 6.
If the measured conductivity in step 4 under stage 2 is not greater than the conductivity
requirements for the pH determined, the water to be examined meets the requirements of the
test for conductivity. If either the measured conductivity is greater than this value or the pH is
outside the range of 5.0-7.0, the water to be examined does not meet the requirements of the
test for conductivity.













































































CHARACTERS

Appearance

Clear and colourless liquid.

TESTS

Nitrates

Maximum 0.2 ppm.

Place 5 mL in a test-tube immersed in iced water, add 0.4 mL of a 100 g/L solution of
potassium chloride R, 0.1 mL of diphenylamine solution R and, dropwise with shaking, 5 mL of
nitrogen-free sulfuric acid R. Transfer the tube to a water-bath at 50 C. After 15 min, any blue
colour in the solution is not more intense than that in a reference solution prepared at the same
time in the same manner using a mixture of 4.5 mL of nitrate-free water R and 0.5 mL of nitrate
standard solution (2 ppm NO
3
) R.

Aluminium (2.4.17)

Maximum 10 ppb, if intended for use in the manufacture of dialysis solutions.

Prescribed solution To 400 mL of the water to be examined add 10 mL of acetate buffer
solution pH 6.0 R and 100 mL of distilled water R.

Reference solution Mix 2 mL of aluminium standard solution (2 ppm Al) R, 10 mL of acetate
buffer solution pH 6.0 R and 98 mL of distilled water R.

Blank solution Mix 10 mL of acetate buffer solution pH 6.0 R and 100 mL of distilled water R.

Bacterial endotoxins (2.6.14)

Less than 0.25 IU/mL.

LABELLING

The label states, where applicable, that the substance is suitable for use in the manufacture of
dialysis solutions.


Ph Eur